Abstract

The application of Preimplantation Genetic Testing (PGT) is no longer limited to conditions presented at birth, but also applied for the adult onset disorders with genetic predisposition, which have never been an indication for prenatal diagnosis. One of such conditions is the inherited predisposition for cardiac disease for which no pre-clinical diagnosis and preventive management exist, with premature or sudden death frequently representing the only clinical realization of the predisposing genes. We have presently performed 75 PGT cycles for 47 couples at risk for producing offspring with predisposition to cardiac diseases, resulting in 37 clinical pregnancies and birth of 32 healthy children, free of genes predisposing to these conditions. This systematic experience demonstrates that PGT may now be a practical means for avoiding the risk of premature or sudden death in the offspring of carriers of a cardiac disease predisposing genes.

Keywords

Introduction

PGT is becoming a useful part of genetic practices and
Assisted Reproductive Technology (ART) [1,2]. In addition
to the traditional indications similar to those in prenatal
diagnosis, it is gradually being applied not only to the disorders
presented at birth, but also to hose that are realized later in life,
such as genetic predisposition to cancer and cardiac disease
[3-5]. A few previous reposts demonstrated the usefulness of
PGT for inherited cardiac disease, allowing the carriers of
predisposing genes to avoid the inheritance of these genes
to their offspring [6-8]. The data showed that avoiding the
inheritance of the predisposing genes to the offspring may
be the only option to prevent the disease, as it may manifest
despite pre-symptomatic diagnosis and follow up. This paper
presents our systematic work on PGT for inherited cardiac
disease, which represents the world’s largest series of 75 PGT
cycles resulting in the birth of healthy children without the
risk of developing an inherited cardiac disease.

Materials and Methods

A total of 75 PGT cycles for 47 couples at risk for
producing an affected progeny with inherited cardiac disease
were performed, which includes 30 cycles for 51 couples
reported earlier [6,7].

To perform PGT the couples at risk underwent a standard
IVF cycle, allowing sampling of material from oocytes and/
or embryos, described in detail previously [8]. Depending
on derivation of predisposing genes from mother or father,
the choice of biopsy materials differed, involving Polar
Body (PB) sampling, in cases of de novo mutations of
maternal origin, and/or embryo biopsy, performed initially by blatomere sampling at the 8-cell stage, or by blsastocyst
biopsy at the present time [8]. The biopsied PBs, blastomeres
or blastocyst samples were tested by the multiplex nested
PCR analysis, directly or after Whole Genome Amplification
(WGA), for the mutations predisposing to cardiac disease and
linked marker analysis in a multiplex heminested system [8].

In cases of advanced reproductive age, 24-chromosome
aneuploidy testing was performed in together with mutation
analysis, using next generation technologies, initially array
CGH and presently NGS (Illumina Inc). Pregnancy outcome
was defined as the presence of a gestational sac with foetal
cardiac activity.

The embryos derived from the embryos free of genetic
predisposition to cardiac disease, based on the mutation
and polymorphic marker information, were pre-selected
for transfer back to patients, while those with predisposing
mutant genes were considered affected , and tested to confirm
the diagnosis.

Results and Discussion

Table 1 summarizes the cardiac disorders for which
PGT was performed, including dilated cardiomyopathy,
determined by 16 predisposing gene mutations (23 cycles for
11 couples), hypertrophic cardiomyopathy, determined by 19
predisposing gene mutations (18 cycles for 12 couples), long
QT, determined by different predisposing gene mutations
(6 cycles for 6 couples), Holt-Oram syndromes determined
by 2 predisposing gene mutations (6 cycles for 3 couples),
ACyl-CoA Dehydrogenase, determined by 5 predisposing
gene mutations (6 cycles for 5 couples) and a few cycles in
couples where cardiac disease was part of other conditions, such as cardioencephalomyopathy (4 cycles for 2 couples),
and Noonan syndrome 1 (6 cycles for 3 couples). All but two
couples tested for cardiac disease were carries of autosomal
dominant mutations, one having no previous affected progeny,
but had a family history of premature or sudden death.

Table 1. List of inherited cardiac diseases and their predisposing gene mutations, for which PGT was performed and the clinical outcome of the procedure (AD ? Autosomal Dominant; AR - Autosomal Recessive; XLR ? X-linked Recessive).

Of 75 PGT cycles performed, 65 resulted in transfer of
91 cardiac disease predisposition free embryos, yielding 37
clinical pregnancies (57% pregnancy rate per transfer) and
birth of 39 predisposition free children. Overall the testing
was done for 65 different predisposing gene mutations,
which was extremely accurate and reliable, with the results
obtained in over 90% of the tested samples, with the
information including also euploidy status. On the basis of
mutation and aneuploidy testing, 91 genetic predisposition
free embryos were transferred in 65 cycles (1.4 mutation
free embryos per transfer cycle on the average). As a result,
37 patients became pregnant after the first or second PGT
cycles and delivered 32 healthy children free of predisposing
gene mutations. No misdiagnosis was observed, suggesting that procedure is highly accurate and acceptable for a wider
clinical application.

Presented results show that PGT may be offered as a
realistic option for couples at high risk for producing offspring
with cardiac disease, to avoid inheritance of the predisposing
genes from parents. It is obvious that information about
such option can be extremely useful for couples at risk, as if
inheritance of these genes is not avoided, their offspring will
be at risk for the cardiac disease development, which may
manifest at the childhood, or later in adult life, with the main
clinical realization of premature or sudden death.

To offer the procedure to those who may actually benefit
from the procedure, family history may appear important as
part of the clinical documentation, including not only cardiac
disease, such of heart attack and sudden death at young ages,
but also information about pacemakers or internal cardiac
defibrillators, arrhythmia or heart surgery in the family
members. No doubt that the penetrance of predisposing
genes depends on the mode of inheritance and the lifestyle, such as an excessive exercise, but there is always risk of
carriers of predisposing genes to contract a cardiac arrest or
sudden death, which is therefore an obvious indication for
PGT. It is also understood that symptoms of inherited cardiac
disease may be easily overlooked, so again the family history
may be the reason for undertaking an exclusion testing of
predisposing gene mutations, which may justify offering
PGT. So it may be expected that cardiac disease predisposing
genes will be an important candidate for preconception
screening program packagers, currently being implemented
in many patient groups, such as in ART practice. With such
a prospective identification of carries of genes predisposing
to inherited cardiac disease, PGT might appear a useful tool
for avoiding the risk for producing offspring with a high
probability of premature or sudden death at their lifespan.